Macrophages are distributed throughout the body where they are poised to detect pathogens and to subsequently alert the immune system to the presence of infection through the production of inflammatory mediators. Inflammatory mediators are critical for pathogen control, but if produced excessively, can result in inflammatory diseases. My long-term goal is to understand the regulation of the production of inflammatory mediators by macrophages during infection. I have identified a novel type of negative regulation of the inflammatory response to pathogens through the TREM-2/DAP12 receptor complex. The specific aims of this proposal seek to define the mechanism of TREM-2 and DAP12 inhibition of inflammatory signaling in macrophages both in vivo and in vitro. The specific aims are: 1. To determine the ligand requirements for TREM-2 and DAP12 inhibition of inflammatory responses in macrophages. I will test the hypothesis that low avidity signals through DAP12 result in inhibition of inflammatory responses, whereas high avidity signals result in activation of inflammatory responses. 2. To determine the mechanism by which TREM-2 and DAP12 inhibit inflammatory signaling in macrophages. These experiments will define how DAP12 signaling results in inhibition of ERK phosphorylation and activation leading to dampening of inflammatory cytokine production in macrophages. 3. To determine why DAP12-deficient mice have enhanced innate immune responses in vivo. I will determine the mechanism for the increased innate response to Listeria monocytogenes infection in DAP12- deficient mice. I also will investigate what myeloid populations are inhibited by DAP12 signaling and whether this correlates with TREM-2 and TREM-2 ligand expression. Results from these studies should provide novel insights into how the early innate immune response to infectious pathogens is controlled. This is a topic of significance in the development of vaccines to prevent infection and immunomodulatory drugs to treat infections with pathogens relevant to biodefense, including Listeria monocytogenes, an Ml AID category B priority pathogen addressed in this proposal.